Methillin resistant S. aureus (MRSA) was first identified in clinical specimen in 1961 - shortly after the introduction of the first beta-Iactamas resistant antibiotic - Celbenin - into clinical practice. The key component of this resistance mechanism - the mecA gene - encoding a protein with very low affinity to all beta-Iactam antibiotics - is not a native gene for S. aureus but has been acquired on multiple occasions from a ''foreign'' source in the form of chromosomal cassettes. MRSA clones have emerged as major causative agents of serious and often life-threatening infections in hospitals worldwide and beginning with the late 1990s MRSA also found its way into the community. The most widely spread epidemic clones of MRSA also acquired resistance traits to the great majority of antimicrobial agents and therapeutic options against such multidrug resistant strains have become reduced to less than a handful of agents. The primary motive of this grant proposal is to find novel intervention strategies against MRSA by exploring more closely the mechanism of beta-Iactam resistance. Critical clues for such novel strategies were observations made in the two collaborating laboratories. The research program will use a combination of genetic, biochemical and metabolomic approaches and will be divided into four Specific Aims. Aim 1 will use metabolomic approaches to identify the mechanisms by which a library of auxiliary mutants can reduce resistance level of an MRSA strain. Aim 2 will compare the very different resistance levels produced by subpopulations of heterogeneously resistant epidemic clones of MRSA. Aim 3 will use genome sequencing to identify determinants in the genetic background of S. aureus that control the level of resistance to oxacillin. Aim 4 will use five contemporary isolates of MRSA recovered in Yr 2011 in New York hospital